A technique for the evaluation of sperm penetrating ability and quality ofbovine semen processed in an extender made with Brackett-Oliphant medium and egg yolk

Egg yolk- sodium citrate (EYC) semen extender was compared with an extender made of Brackett-Oliphant medium and egg yolk (BOEY). Ejaculates were divi ded into equal portions, processed and frozen. Semen was thawed and evaluat ed for quality. Additional semen was thawed, stained with Hoechst 33342 and the spermatozoa capacitated, after which they were coincubated with zone-f ree hamster oocytes to determine their penetrating ability. Sperm penetrati on of non-compressed, unfixed oocytes was evaluated using an optical sectio ning technique on a standard research microscope. Sperm penetration was con sidered successful if a fluorescing sperm head was observed within the livi ng oocyte in a hanging drop of fertilization medium. There were small diffe rences in percentage of secondary abnormalities and percentage of progressi ve motility immediately after thawing between spermatozoa extended in EYC o r BOEY diluent. There were no differences due to by extender composition in percentage of spermatozoa with intact acrosomes or percent of progressivel y motile after a 3 h incubation at 37 degrees C, nor the percentage of sper matozoa with head abnormalities. While there were significant correlations between all seminal quality characteristics, no quality measurements were c orrelated to percentage of oocyte penetration. The new penetration evaluati on method allowed for examination of the fertilized oocytes using fluoresce nt microscopy initially and again after re-incubation for further developme nt. (C) 1999 by Elsevier Science Inc.